Transmissible spongiform encephalopathies (TSEs), also known as prion diseases, are a group of neurodegenerative diseases affecting a wide variety of mammals including sheep and goats (scrapie), cervid spp. (chronic wasting disease), and humans (Creutzfeldt-Jakob disease). A central event in TSE disease involves the conversion of the normal host cellular prion protein (PrPC) to a partially protease-resistant, aggregated, disease-associated isoform (PrPSc). TSE-induced pathology is usually associated with PrP-res deposition, but the mechanism of neurodegeneration is not understood. Like prion diseases, other neurodegenerative diseases involve the deposition of pathological protein aggregates including Alzheimers and Parkinsons disease. Collectively, these diseases are termed protein misfolding diseases because they are associated with the accumulation of misfolded host proteins. Critical processes for transmission of prions between and within hosts include neuroinvasion and intercellular spread of PrPSc. The membrane association of PrP via a glycosylphosphatidylinositol (GPI)-anchor may have an important role in modulating these processes. Our work in this project is focused on elucidating the role of GPI and other forms of membrane anchoring in modulating the aggregation and intercellular spread of aggregation-prone proteins. In 2013, we have continued experiments to investigate how membrane anchoring allows the propagation of protein aggregates as prions and the effects of membrane anchoring on the structure of self-propagating protein aggregates. One line of investigation involves the characterization of the structure of membrane-bound aggregates of a GPI-anchored protein called Sup35NM-GPI. Without GPI-anchoring, Sup35NM assembles into amyloid fibrils. However, our data visualizing aggregate structure by immunogold microscopy and staining with several sensitive amyloid stains suggest that GPI-anchoring directs Sup35NM assembly into non-amyloid membrane-bound aggregates.